Characterization of the biology and pathology of an arbovirus-insect host interaction during persistence is the overall goal of this research program. Virus and host cell responses to acute and chronic infections will be examined following oral inoculation of adult female mosquitoes with the prototype Alphavirus, Sindbis virus (SIN). Specific Aims include I: Selection for a colony of Ae. albopictus mosquitoes efficient at blood feeding on an artificial membrane, II: Analysis of SIN-associated pathology in the adult mosquito, III: Selection of persistence variants of SIN following chronic infection in female mosquitoes and IV: Comparison between Aedine and Culex responses to SIN. SVHR and the cDNA clone, pTR339 a molecular construct retaining the consensus sequence of AR339, will be used to produce wild type virus. Infectious blood meals will be proffered to Ae. albopictus, Ae. aegypti and Culex quinquefasciatus mosquitoes via an artificial membrane apparatus and females that engorge will be sampled at day 5 p.i. and day 30 p.i. representing acute and chronic infections, respectively. Changes seen in both the invertebrate host and the virus during persistence in the insect milieu will be examined. Our lab has documented SIN-associated pathology in mosquito salivary glands and midgut-associated muscles, and we will investigate the involvement of apoptosis as a mechanism of virus-associated pathology. Natural virus variants, selected for during chronic infection will be isolated; persistent (SIN-PI#), tracheole (SIN-TRA#) & saliva (SIN-SPIT#). Strategies to select for tissue-specific genetic variants will increase our knowledge of pathogen transmission and maintenance of arboviruses in nature. The current global-climate of emerging and re-emerging diseases supports investigations into the holistic biology of arbovirus-mosquito interactions, an essential link in the chain of pathogen transmission, a link, which if broken would prove invaluable in the control of the spread of these diseases. A mosquito host remains persistently infected with an arthropod-borne-virus for the lifespan of the insect, a situation which increases the chances of virus transmission. We seek to understand the biology and pathology associated with viruses isolated from chronically infected mosquitoes. This research is relevant to public health because investigations into the cellular and molecular aspects of virus persistence in the insect host will further the prevention of mosquito-borne infectious diseases, in the current global-climate of emerging and re-emerging diseases. [unreadable] [unreadable] [unreadable]